Human adult tissue-specific stem cells have clinical utility due to their ability to repair and/or replace damaged tissue. However, identification of adult stem cells has proven to be difficult, because of a lack of proper tissue-specific stem cell markers. Limiting their clinical application further, they have a finite lifespan in culture and restricted differentiation capacity, especially compared to human embryonic stem cells (ESCs). Among adult stem cells that have been isolated thus far, bone marrow-derived mesenchymal stem cells (BM-MSCs) are the most well characterized. The BM-MSCs were identified over 10 years ago and give rise to various differentiated cell types of mesodermal origin. However, isolation of BM-MSCs is quite painful for patients and once isolated, they are difficult to maintain in culture because they reach senescence rapidly (usually by 8 passages), lose their differentiation capacity rapidly, and, on some occasions, become neoplastic after extended in vitro culture. Other source of stem cells include dental pulp, Wharton's jelly, amniotic membrane, and adipose tissue, yet all of them have limited lifespan and differentiation capabilities.
Among the specific stem cell markers, CD34 is found on early hematopoietic and vascular-associated tissues. CD34 is a 116-kD type I transmembrane glycophosphoprotein: however, little is known about its exact function. In the hematopoietic system, cells expressing CD34 on the surface can, upon cytokine or growth factor stimulation, expand and differentiate into all the lymphohematopoietic lineages. Thus, CD34 has been used as a marker to assist in the identification and isolation of lymphohematopoietic stem/progenitor cell populations; more recently, it has been employed as a marker to help identify other tissue-specific stem cells, including muscle satellite cells and epidermal precursors. Recently, it has found that CD34-positive stromal cells are distributed in various organs including breast, fallopian tubes, thyroid gland, colon, pancreas, uterine cervix, and testis (Kim J, Seandel M, Falciatori I, et al. CD34+ testicular stromal cells support long-term expansion of embryonic and adult stem and progenitor cells. Stem Cells. 2008; 26:2516-2522). In adipose-derived stromal cell (ASC) populations, CD34-positive cells are resident pericytes that play a role in vascular stabilization by mutual structural and functional interactions with endothelial cells. In another study, CD34-positive cells showed higher proliferative and colony-forming capacity than CD34-negative cells, but lower differentiating capability. Thus, the authors suggested that CD34 expression was inversely correlated to the physiological process of differentiation from an immature status into specific lineages (Suga H, Matsumoto D, Eto H, et al. Functional implications of CD34 expression in human adipose-derived stem/progenitor cells. Stem Cells Dev. 2009; 18:1201-1210). However, CD73 is a glycosyl phosphatidylinositol (GPI)-linked, membrane-bound glycoprotein that hydrolyzes extracellular nucleoside monophosphates into bioactive nucleoside intermediates. This antigen is found in most cell types including mesenchymal stem cells, subsets of B-cells and T-cells and endothelial cells. Thus, this molecule has been used as a maker to identify MSCs originating from several different tissues. Interestingly, almost none of the MSCs isolated so far show both CD73 and CD34 expression.
Meanwhile, mammalian testis consists of germ cells and various types of somatic cells. The lack of specific markers has made it difficult to identify and localize potential stem cells in tissues. Several researchers have isolated and propagated unipotent stem cells such as spermatogonial stem cells (SSCs) and Leydig stem cells (Kanatsu-Shinohara M, Ogonuki N, Inoue K, et al. Long-term proliferation in culture and germline transmission of mouse male germline stem cells. Biol Reprod. 2003; 69:612-616; and Ge R S, Dong Q, Sottas C M, et al. In search of rat stem Leydig cells: identification, isolation, and lineage-specific development. Proc Natl Acad Sci USA. 2006; 103:2719-2724). In addition, primordial germ cell-derived embryonic stem cell-like cells have been generated using testis biopsies from both human and mouse (Conrad S, Renninger M, Hennenlotter J, et al. Generation of pluripotent stem cells from adult human testis. Nature. 2008; 456:344-349; Guan K, Nayernia K, Maier L S, et al. Pluripotency of spermatogonial stem cells from adult mouse testis. Nature. 2006; 440:1199-1203; Seandel M, James D, Shmelkov S V, et al. Generation of functional multipotent adult stem cells from GPR125+ germline progenitors. Nature. 2007; 449:346-350; Kanatsu-Shinohara M, Inoue K, Lee J, et al. Generation of pluripotent stem cells from neonatal mouse testis. Cell. 2004; 119:1001-1012). These cells differentiated into cells of all 3 germ layers and formed tumors when they were injected into NOD-SCID mice (Conrad S, Renninger M, Hennenlotter J, et al. Generation of pluripotent stem cells from adult human testis. Nature. 2008; 456:344-349).
However, studies on testis somatic stem cells have been rare. Only recently was a MSC-like population isolated in adult human testes and partially characterized by differentiating it into mesodermal-lineage cells (Gonzalez R, Griparic L, Vargas V, et al. A putative mesenchymal stem cells population isolated from adult human testes. Biochem Biophys Res Commun. 2009; 385:570-575). These cells were positive for CD90 and negative for CD34, suggesting they were testis-derived MSCs with limited lifespans in vitro. In the mouse, CD34-positive stromal cells efficiently support the proliferation of adult spermatogonial progenitor cells (Seandel M, James D, Shmelkov S V, et al. Generation of functional multipotent adult stem cells from GPR125+ germline progenitors. Nature. 2007; 449:346-350). However, no study has been carried out to determine if CD34/CD73-double-positive testis stromal cells are another somatic stem cell source and, if so, what their differentiation and proliferation capabilities are.